The long-term objective of this project is to understand the signaling mechanisms and physiological functions of the newly discovered Rac-cGMP pathway. Many membrane-signaling receptors can increase the cellular levels of the ubiquitous second messenger cyclic GMP (cGMP). However, the molecular mechanism by which these membrane-signaling receptors increased cGMP was not known. We recently uncovered a new Rac-cGMP signaling pathway. The small G protein Rac uses its effector PAK (p21-activated kinase) to allosterically activate transmembrane guanylyl cyclases (GCs). These findings reveal a general mechanism for diverse signaling receptors to modulate physiological responses through cGMP. Although the kinase activity of PAK is required, PAK does not directly phosphorylate GCs. Instead, autophosphorylation of PAK is needed to maintain its activated configuration. The active form of PAK then allosterically activates transmembrane GCs. The main focus of this application is on the biochemical mechanism by which Rac, through its effector PAK, regulates the activity of transmembrane guanylyl cyclases. We will investigate the allosteric activation of transmembrane guanylyl cyclases by PAK in the Specific Aim 1. In the Specific Aim 2, we will explore the signaling molecules downstream of cGMP. We will investigate the physiological function of the Rac-cGMP pathway in the Specific Aim 3. PUBLIC HEALTH RELEVANCE: This research is directly related to human health. Both Rac GTPase and the second messenger cGMP are critical regulators of many physiological functions. Cell migration and vascular permeability are essential for vascular development and are involved in cardiovascular diseases. Investigations of the signaling mechanisms and physiological functions of the Rac-cGMP pathway will significantly advance our understanding of cardiovascular function and diseases.